The developed sensor also showed a sensitive and selective response towards DENV 2 E-proteins compared to DENV 1 E-proteins and ZIKV (Zika virus) E-proteins. Overall, it was concluded that the Au/DSU/amine-functionalized rGO–PAMAM thin film-based SPR sensor has potential to serve as a rapid clinical diagnostic tool for DENV infection.Dengue viral infection is one of the most common deadliest diseases and has become a recurrent issue for public health in tropical countries. Although the spectrum of clinical diagnosis and treatment have recently been established, the efficient and rapid detection of dengue virus (DENV) during viremia and the early febrile phase is still a great challenge. In this study, a dithiobis /amine-functionalized reduced graphene oxide-–polyamidoamine dendrimer  thin film-based surface plasmon resonance (SPR) sensor was developed for the detection of DENV 2 E-proteins. Different concentrations of DENV 2 E-proteins were successfully tested by the developed SPR sensor-based system. The performance of the developed sensor showed increased shift in the SPR angle, narrow full-width–half-maximum of the SPR curve, high detection accuracy, excellent figure of merit and signal-to-noise ratio, good sensitivity values in the range of 0.08–0.5 pM (S = 0.2576°/pM, R The demand for rapid, sensitive, and quantifiable methods for the detection of dengue virus is significant. Optical sensors have emerged as a promising technology to potentially identify biological phenomena. They have the advantages of being highly sensitivity, allowing fast detection and real-time measurements, and having a simple configuration ,4,5,6,7.2) renders it hydrophilic and increases its interfacial binding to materials of interest, thus making it more adaptable as a sensing platform for the detection of dengue virus [Graphene oxide (GO)-based sensors have received great attention due to their exciting properties including abundant functional groups, low thickness, very low mass, large specific area, high π-conjugation structure, and high mechanical strength ,26,27,28ue virus ,35,36. Lue virus ,38,39,40With no effective and licensed vaccines, early detection of DENV is crucial to prevent severe clinical complications, which can cause failure of the circulatory system and the liver and death if not adequately managed. There are four distinct serotypes of dengue viruses, i.e., DENV 1 to DENV 4, and each serotype consists 3 structural proteins  protein) and 7 non-structural proteins ,45,46,47Herein, we developed an SPR sensor based on an amine-functionalized rGO–PAMAM composite, with monoclonal antibodies immobilized on self-assembled dithiobis  to detect and quantify the dengue virus. The sensing performances of the proposed sensor, i.e., linear sensitivity, detection accuracy, full-width half maximum, signal-to-noise ratio, figure of merit, selectivity, and matrix effect are discussed in detail. To the best of our knowledge, this is the first report of the detection of DENV at the low concentration of 0.08 pM in 8 min using an Au/DSU/amine-functionalized rGO–PAMAM/IgM thin film-based SPR optical sensor.N-Ethyl-N-(3-(dimethylaminopropyl) carbodiimide (EDC) was bought from Fluka, Switzerland. The standard powder of recombinant DENV 2 E-proteins and antibodies (IgM) against dengue type 2 envelope proteins were purchased from Meridian Life Science. The preparation of amine-functionalized rGO was begun by amalgamating the graphene oxide with EDC for 5 min, followed by the addition of EDA. After the solution was vigorously stirred, the suspension was slowly dissolved and started to change its color from brown to dark black. Then, the suspension was washed for 5 times with ethanol and centrifuged to remove excessive EDA and EDC and then was allowed to dry in an oven at 60 °C for at least 1 h. Afterward, the PAMAM solution with the desired concentration was mixed to the amine-functionalized rGO to obtain a composite. Unless otherwise indicated, all antibodies and antigen solutions were diluted in 10 mM phosphate-buffered saline (PBS) at pH 7.4. All chemicals were of reagent or higher grade, and deionized water was used throughout the experiments.DSU (MW = 628.84 g/mol) was purchased from Dojindo Japan. Graphene oxide was purchased from Graphanea, Spain. PAMAM dendrimer , ethylenediamine (EDA), N-hydroxysuccinimide (NHS), bovine serum albumin (BSA) were purchased from Sigma Aldrich, Germany. To produce the best SPR peak for sensor application, a metal deposition technique to obtain a thin Au film (~50 nm) with high thickness tolerance is required. The generally used methods for metal thin-film deposition are electroplating, sputtering, and evaporation  ,55,56. SFor the detection of DENV 2 E-proteins, the sensor film was mounted on a prism with a refractive-index-matching liquid and loaded onto the SPR rotating stage for SPR measurements. θ, versus DENV 2 E-protein concentrations, as follows:There are several parameters, i.e., sensitivity, full width at half minimum (FWHM), detection accuracy (DA), signal-to-noise ratio (SNR), figure of merit (FOM), selectivity, and spike recovery that are used to analyze the performance of SPR sensors. The sensitivity of an SPR sensor can be defined by calculating the slope in the graph of the resonance angle shift, ΔA second parameter is the angular width of the SPR curve for the half value of the maximum reflectance, which is known as the FWH). The value of FWHM should be as small as possible in order to get an accurate reading on the dip of the SPR curve . TherefoTo comprehensively evaluate the performance of the SPR signal, the SNR can be determined according toFinally, the FOM can be defined as the ratio between the spectral sensitivity and the width of the SPR curve and can be written asThe surface morphology and roughness of the sensor films were imaged using an atomic force microscope  in Scan Asyst mode. This measurement was performed in ambient atmosphere at room temperature.θ) was determined from the difference between the resonance angle of the antigen and the resonance angle of the reference solution. It was found that increases of Δθ of 0.0914°, 0.0999°, 0.1708°, 0.1866°, and 0.1945° were obtained when detecting 0.08 pM, 0.1 pM, 0.3 pM, 0.5 pM, and 1 pM DENV 2 E-proteins, respectively. These Δθ can be attributed to the changes in the refractive index of the sensor surface which in turn changed the real part of the dielectric constant of the gold film caused by the binding of DENV 2 E-proteins. It was inferred that a change of the thickness of the sensing layer would also result in a slight angle shift of SPR, as the evanescent wave possesses longer penetration depths [θ varied with time when detecting different concentrations of DENV 2 E-proteins. It can be seen that the proposed sensor achieved a stable detection time of 8 min for the lowest concentration of DENV 2 E-proteins. Therefore, all DENV E-proteins concentrations were left for 8 min before the SPR curve was recorded.n depths ,59. The −1 was achieved in the detection of 0.00008 nM DENV E-proteins.θmax is the maximum SPR shift at saturation, C is the concentration of DENV, and DK is the equilibrium dissociation constant. Typically, the strength of biomolecular interactions is quantified through DK . The fitted graph gave a DK of 0.1308 pM with R2 of 0.97, which was found to be consistent with the standard DK value for protein interactions (DK < 10 nM) [DK value led to higher binding affinity of the ligand for its target. The apparent AK values for the binding of DENV 2 E-proteins to the sensor surface was 7.6452 TM−1, indicating the high potential for the use of the proposed SPR sensor in detecting DENV 2 E-proteins.To investigate the binding between Au/DSU/amine-functionalized rGO–PAMAM/IgM-based SPR sensor and target, the changes in the resonance angle as a function of the concentration of DENV 2 E-proteins from 0.08 to 1 pM were plotted and fitted according to the Langmuir adsorption model , represe< 10 nM) ,66. A sm2, of 0.92. The slope of the linear fit was considered to be the sensitivity value of the proposed SPR sensor, thus yielding a sensitivity of 0.2576°/pM. It appears that DENV 2 E-proteins can be sensitively detected at the lowest concentration of 0.08 pM. This is possible thanks to the great penetration depth of the SPR evanescent field along the DSU/amine-functionalized rGO–PAMAM/IgM sensor layer.In order to determine the sensitivity of the proposed sensor film for DENV 2 E-proteins, a linear correlation curve was plotted and is presented in To determine the selectivity of the Au/DSU/amine-functionalized rGO–PAMAM/IgM sensor film towards DENV 2 E-proteins, other envelope proteins, such as DENV 1 E-proteins and ZIKV E-proteins, at the concentration of 10 pM were tested. An excellent sensor should be highly selective in the detection of DENV 2 E-proteins. The results are shown in AFM images were taken before a and aft−1. The highest FOM and DA values were obtained when detecting 0.08 pM of DENV 2 E-proteins and corresponded to the minimum values of SNR. The selectivity of the proposed sensor was demonstrated, since the SPR response to other proteins was reduced compared to that to DENV 2 E-proteins. It can be concluded that the developed SPR sensor has a great potentiality for rapid DENV diagnostic analyses or point-of-care tests.In this study, a DSU/amine-functionalized rGO–PAMAM thin film-based SPR sensor was developed for the detection and quantification of DENV 2 E-proteins. The results demonstrate that the developed sensor could successfully monitor the changes in the SPR angle when detecting the lowest concentration of DENV 2 E-proteins, i.e., 0.08 pM, within 8 min. The sensitivity and binding affinity constant of the developed sensor were 0.2576°/pM and 7.6452 TM


The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has collapsed health systems worldwide. In adults, the virus causes severe acute respiratory distress syndrome (ARDS), while in children the disease seems to be milder, although a severe multisystem inflammatory syndrome (MIS-C) has been described. The aim was to describe and compare the characteristics of the severe COVID-19 disease in adults and children.This prospective observational cohort study included the young adults and children infected with SARS-CoV-2 between March–June 2020 and admitted to the paediatric intensive care unit. The two populations were analysed and compared focusing on their clinical and analytical characteristics and outcomes.Twenty patients were included. There were 16 adults (80%) and 4 children (20%). No mortality was recorded.n = 7, 43.7%). Thirteen (81.3%) needed mechanical ventilation, with a median PEEP of 13 (IQR 10.5–14.5). Six (37.5%) needed inotropic support due to the sedation. Eight (50%) developed a healthcare-associated infection, the most frequent of which was central line-associated bloodstream infection . One patient developed a partial pulmonary thromboembolism, despite him being treated with heparin.All the adults were admitted due to ARDS. The median age was 32 years (IQR 23.3–41.5) and the most relevant previous pathology was obesity (All the children were admitted due to MIS-C. Two (50%) required mechanical ventilation. All needed inotropic support, with a median vasoactive-inotropic score of 27.5 (IQR 17.5–30).p < 0.001). The biomarker values were higher in children than in adults: mid-regional pro-adrenomedullin 1.72 vs. 0.78 nmol/L (p = 0.017), procalcitonin 5.7 vs. 0.19 ng/mL (p = 0.023), and C-reactive protein 328.2 vs. 146.9 mg/L (p = 0.005). N-terminal pro-B-type natriuretic peptide and troponins were higher in children than in adults .The difference in the inotropic requirements between the two populations was statistically significant (37.5% vs. 100%, Adults and children had different clinical manifestations. Adults developed severe ARDS requiring increased respiratory support, whereas children presented MIS-C with greater inotropic requirements. Biomarkers could be helpful in identifying susceptible patients, since they might change depending on the clinical features. The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which first appeared in December 2019 in Wuhan, has caused the coronavirus (COVID-19) related pandemic and has been declared a significant threat to international health by the World Health Organization. As of 20 December 2020, 75 million cases have been reported across the globe, causing more than 1,6 million deaths [Interestingly, not all patients have the same manifestations; while some of them have hypoxemia due to pneumonia with a ventilation/perfusion mismatch (pulmonary vasoconstriction), others have this due to lower compliance and severe ARDS . By contBoth ARDS and MIS-C are most likely post-infectious manifestations. Little is known about the specific immunity response after a SARS-CoV-2 infection. It seems that there is an initial peak viral load , and then this declines as antibodies (IgM and IgG) increase within days to weeks of symptom onset, just when inflammatory signs appear . HoweverSince the clinical presentation of the severe forms of the disease implies a cytokine release syndrome with an increase in interleukin-6 (IL-6), tocilizumab has been pointed to as a useful treatment to decrease the inflammatory response . In chilDuring the COVID-19 pandemic, both adult and paediatric patients were admitted to our paediatric intensive care unit (ICU), and a different clinical presentation and evolution were detected in each group. The aim of this study was to describe the characteristics of the disease in each specific population and to analyse the differences between adults and children regarding the clinical manifestations and analytical data.This was a prospective and observational cohort study, performed in the paediatric ICU of Hospital Sant Joan de Déu in Barcelona, which is a paediatric tertiary referral hospital. This paediatric ICU has 28 ventilator-beds and sees 1200 admissions per year. All the patients infected with SARS-CoV-2  admitted to the paediatric ICU between March and June 2020 were included, both adults and children. These two populations were analysed separately in order to compare their clinical characteristics and outcomes.The following demographic data were collected: age, gender, comorbidities, Paediatric Risk of Mortality Score (PRISM III) at admission (11), and reason for admission (respiratory or cardiovascular failure). Respiratory support was also analysed: positive end-expiratory pressure (PEEP), ventilator and oxygenation requirements , and length of mechanical ventilation (MV) and non-invasive ventilation (NIV). Haemodynamic support requirements were analysed using the vasoactive inotropic score (VIS) (11) and the length of the inotropic treatment. The need for heparin treatment was analysed: a high heparin dose was considered as 1 mg/kg/12 h and a low heparin dose was considered as 0.5 mg/kg/12 h. The microbiological data collected included microorganisms isolated in the blood, respiratory tract, and urine cultures. Antibiotic requirements and lengths of treatment were recorded. The analytical biomarkers recorded were C-reactive protein, procalcitonin, mid-regional pro-adrenomedullin (MR-proADM), ferritin, D-dimer, lactate dehydrogenase, and blood counts. N-terminal pro-B-type natriuretic peptide (NT-proBNP) and troponins were analysed in order to assess cardiac dysfunction.Serum levels of procalcitonin and MR-proADM were determined by immunofluorescense by means of a Time-Resolved Amplified Cryptate Emission (TRACE) and a Kryptor analyser . C-reactive protein was obtained using the immunoturbidimetric assay (COBAS INTEGRA ROCHE).The length of stay in the hospital and in the paediatric ICU, as well as the mortality rate, were also recorded.ARDS, identified following the Berlin Definition, specifically, respiratory failure with bilateral pulmonary opacities with PaO2 < 300 mmHg and PEEP or CPAP > 5 cmH2O [ 5 cmH2O .MIS-C in children and adolescents, identified according to the preliminary definition provided by the World Health Organization, which specifies the presence of shock, with elevated biomarkers and a positive COVID-19 test, without other microbial causes of inflammation [ammation . These pammation .The patients were classified, depending on their clinical presentation, as having:A statistical analysis was performed using the IBM SPSS 25.0 Statistics® programme. Categorical variables were indicated as frequency (n) and percentage (%), whereas continuous variables were summarised as median and interquartile range (IQR) because they did not follow a normal distribution, according to the Shapiro-Wilk test. The comparison of categorical variables was performed using the χ2-test or Fisher’s exact test. Continuous variables were compared using the Mann-Whitney U test. Probability values of less than 0.05 were considered statistically significant.During the study, 20 patients were included. There were 16 adults (80%) and 4 children (20%). All the patients had a positive PCR result from the nasopharyngeal sample except one child, who had suggestive clinical features and a positive IgG. No patient needed extracorporeal membrane oxygenation support and no deaths were recorded.n = 7, 43.7%). Among them, 13 (81.3%) needed MV, with a median maximum PEEP of 13 (IQR 10.5–14.5). Prone position was required in 10 patients (76.9%) and recruitment manoeuvres in 5 patients (38.5%). Six patients (37.5%) required inotropic support, in all cases due to sedation. All the patients received treatment with heparin and 6 of them (37.5%) were on high doses. The specific treatment related to COVID-19 is detailed in Table The 16 adults all presented with ARDS. The median age was 32 years (IQR 23.3–41.5), and 11 (68.8%) had a relevant previous pathology, the most frequent of which was obesity  The microbiological details are summarised in Table Eight patients (50%) developed a healthcare-associated infection, and the most frequent of them were central line-associated bloodstream infections , for procalcitonin it was 0.19 ng/mL (IQR 0.11–0.37), and for C-reactive protein it was 146.95 mg/L (IQR 57.6–193). The detailed results of the blood sample tests are included in Table The adult patients had a median length of stay in the ICU of 12 days (IQR 5.7–17), and they had a total length of stay in the hospital of 21 days (IQR 13.7–29.8).Four children were admitted during the study period. None of them had a pre-existing disease or risk factor. All presented with MIS-C and required admission to the paediatric ICU due to shock. Two of them (50%) required MV, with low ventilator parameters, and two (50%) required NIV. All the patients needed inotropic support due to the shock, with a median VIS of 27.5 IQR 17.5–30) and a median length of inotropic treatment of 3 days (IQR 2–4.7). Detailed information on the respiratory and haemodynamic support is provided in Table 7.5–30 anOn the blood test, the median lymphocyte count was low, as well as the prothrombin time. IL-6 was high at admission. The median for MR-proADM was 1.72 nmol/L (IQR 1.55–2.77), for procalcitonin, 5.7 ng/mL (IQR 1.3–7.77), and for C-reactive protein, 328.2 mg/L (IQR 252.8–351.15). The median values of the NT-proBNP and troponins were elevated in paediatric patients.The results of the blood tests are summarised in Table All the patients received antibiotic treatment. None of them developed a healthcare-associated infection.One patient developed coronary aneurisms. During their stay in the paediatric ICU, echocardiograms were performed routinely and showed systolic dysfunction secondary to the shock, but no aneurisms were detected. The coronary aneurisms appeared during the follow-up period, while the patient was in the hospitalisation ward.The paediatric patients had a median length of stay in the ICU of 6 days (IQR 4.3–7), and they had a total length of stay in the hospital of 12 days (IQR 11–13).p < 0.001). Children had greater inotropic treatment requirements due to the shock, while adults needed it due to the sedation and required lower doses; the differences in the reason for administering inotropic treatment were statistically significant (p = 0.005).While adults were admitted due to ARDS, paediatric patients required admission to the paediatric ICU due to MIS-C (p = 0.006). Children presented higher values of procalcitonin, C-reactive protein, and MR-proADM .There were differences in the blood tests. Children had lower prothrombin time in comparison with adults .NT-proBNP and troponins were higher in children than in adult patients . This is relevant, because most of them had no other risk factors. It has already been reported that obesity might be a risk factor for respiratory diseases, as obesity is associated with a decreased expiratory reserve volume, functional capacity, and compliance. Moreover, the inflammatory cytokines associated with obesity might worsen the evolution of these patients when they contract COVID-19 infections . Accordi2/FiO2 ratio is < 200), and recruitment manoeuvres . This strategy was successful, because the evolution of the patients was good overall, except for one partial pulmonary thromboembolism. This patient was already out of the ICU when it happened and was on a high-dose heparin treatment regime (1 mg/kg/12 h). He needed 2 days of NIV support, without the need for inotropic agents, and his clinical evolution was good afterwards. COVID-19 patients have several risk factors for developing pulmonary thromboembolism. The most important one seems to be the presence of an inflammatory state that leads to hypercoagulability, but they also have other risk factors common in critically ill patients, like prolonged immobilisation [The respiratory treatment of patients with ARDS that needed MV were treated according to the current recommendations for respiratory support : high PElisation . An elevlisation , 19. Heplisation .The respiratory support provided to children was only due to haemodynamic reasons. This is because the clinical features observed in children were completely different. The main problem was shock, not respiratory repercussions. They all needed high doses of inotropic agents, which was a factor that distinguishes them from the adult population. This is consistent with the results of the paediatric study performed by Ramcharan et al., in which all patients had some degree of cardiac dysfunction and their inotropic support requirements were high . In lineTaking into account other parametres on the blood test, the paediatric patients showed coagulopathy and an increase in ferritin, lactate dehydrogenase, and D-dimer. There are few reports about critically ill children with COVID-19, but some of them have described similar features, with cardiogenic shock and elevated inflammatory parameters on the blood test .Both paediatric and adult patients had high levels of IL-6. This is consistent with the IL-6 signalling pathway, which seems to be involved in the damage produced by COVID-19. Tocilizumab, which specifically binds soluble and membrane-bound IL-6 receptors, inhibits the binding of IL-6 with its receptor, and consequently inhibits the signal transduction; therefore, it can prevent cytokine release syndrome in this post-infectious inflammatory situation . We woulIn this sample, no mortality was recorded, neither in the adults nor in the paediatric cases. In other articles, high mortality rates have been reported (some as high as 61.5% of the patients admitted to the ICU); in most cases, this was related to multiorgan failure and occurred in elderly patients . It is tWe acknowledge several limitations in this study. On the one hand, the sample is relatively small, and on the other hand, the adults are young, so the results might not be able to be extrapolated to other populations. However, we think that it provides valuable information, because this is one of the first studies including both children and young adults in critical condition.To sum up, in this sample we observed two completely different entities: adults with ARDS and children with MIS-C. While in the first cohort the main problem was respiratory, with ARDS and high respiratory support requirements, in the second group the problem was haemodynamic in nature, requiring inotropic support to improve the situation. We believe that this could be explained by the fact that the inflammatory response to SARS-CoV-2 seems to be different in adult and paediatric patients, being localised in the lungs or generalised in a multisystem syndrome, respectively. However, this is a small sample, thus further studies with larger samples should be performed in order to confirm these findings.


Selective laser sintering (SLS) is being developed for dental applications. This study aimed to investigate the properties of Ti-6Al-4V and pure titanium specimens fabricated using the SLS process and compare them with casting specimens. Besides, the effect of the building direction on the properties of the SLS specimens was also investigated. Specimens were prepared by SLS using Ti-6Al-4V powder or pure titanium powder. Casting specimens were also prepared using Ti-6Al-4V alloys and pure titanium. The mechanical properties (tensile strength and elongation), physical properties ; corrosion resistors (color difference and corrosion), and surface properties  were examined. Both Ti-6Al-4V and pure titanium specimens produced using the SLS process had comparable or superior properties compared with casting specimens. In comparing the building directions, specimens fabricated horizontally to the printing platform showed the greatest tensile strength, and the surface roughness scanned in the horizontal direction to the platform showed the smallest. However, there was no significant effect on other properties. Thus, the SLS process with Ti-6Al-4V powder and pure titanium powder has great performance for the fabrication of dental prosthesis, and there is a possibility for it to take the place of conventional methods. In recent years, additive manufacturing (AM) has been widely used in various fields and has been developed as a new molding technique . This teSeveral additive manufacturing processes are being applied in dentistry to manufacture casting patterns, provisional restorations, surgical guides, or dental models ,5,6,7,8;Recently, titanium materials have been developed as powders for the AM process. There are several reports showing that frameworks and clasps of removable dentures and fixed dentures can be produced by AM using titanium materials ,14,15,16Therefore, this study aimed to clarify the properties of specimens molded by AM using titanium alloy (Ti-6Al-4V) powder and pure titanium powder. In addition, we investigated the effect of the building direction on the properties of the specimens. Finally, we compared the properties of the AM specimens with those of specimens cast with titanium alloy and pure titanium to examine the applicability of AM with titanium materials.The metal materials used in this study are listed in The DU was molded to set the angle formed by the building direction and major axis at 0°, 45°, and 90° . For theAs comparative controls, specimens with the same size as above were prepared by casting using the metals shown in These casting patterns were invested using a titanium-exclusive investing material . Casting was prepared using a titanium-exclusive casting machine . Ti-6Al-4V casting was performed at a mold temperature of 430 °C and casting temperature of 1540–1650 °C; pure Ti casting was performed in an argon atmosphere at a mold temperature of 430 °C and casting temperature of 1668 °C, respectively. Post-processing was performed in the same manner as the specimens produced by SLS process.In the tensile test, the tensile strength and the elongation to time of fracture (matching method) of the DU specimens were measured. This test was performed at a crosshead speed of 1.0 mm/min using a universal testing machine . The SLS-0, SLS-45, SLS-90, and casting specimens were subjected to this test, and the influence of the building direction of the specimens on the mechanical properties was investigated.The surface roughness, contact angle to water, and Vickers hardness were measured in a physical property test using BL specimens. Surface roughness was measured using a surface roughness measuring instrument . The contact angle was measured with a contact angle meter . The Vickers hardness was measured with a Vickers hardness tester  at a measurement load of 50 kgf and a load holding time of 15 s. These measurements were performed on the surface condition as sintered. Next, the specimen surface was polished to a mirror-like finish, and the same measurements were performed on the surface after polishing. The properties of the specimen surface before (as sintered) and after polishing were compared. These tests were performed on the Z plane (SLS-Z), XY plane (SLS-XY), and the cast specimen, and the effect of the building direction of the specimen on the physical properties was investigated.Two types of immersion tests were performed: a discoloration test (using sodium sulfide and lactic acid solutions) and a corrosion test (using lactic acid solution). For these tests, the BL was cut into a plate of 10 × 10 × 1 mm using a precision cutting machine . The specimen cut vertical to the building stage  is referred to as SLS-Z, and the specimen cut horizontal to the building stage  is referred to as SLS-XY. The two immersion tests were performed on SLS-Z, SLS-XY, and cast specimens, and the effects of the building direction of the specimen on discoloration and corrosion were investigated.The plate specimen surface was polished with water-resistant abrasive paper ). Subsequently, the color, surface area, and weight of the specimen were measured (surface area and weight were measured only for the lactic acid immersed specimen). Color was measured with a colorimeter . The surface area was measured with a digital microscope . Weight was measured with a precision balance . Sodium sulfide solution  or lactic acid solution (10 g/L) was placed in a glass container (capacity approximately 150 mL). One specimen was immersed in the solution so that it did not contact the bottom of the container, and left at 37 °C for three (sodium sulfide solution) or seven days (lactic acid solution) according to ISO 10271:2011 . After iIn the discoloration test, the color difference (ΔE*ab) was calculated from chromaticity coordinates of the specimen measured before and after immersion. To relate the color change measured by the spectrophotometer to a clinical environment, the values of ΔE* ab were converted to National Bureau of Standards units (NBS units) through the equation, NBS units = ΔE*ab × 0.92.In the corrosion test, weight loss due to corrosion (lactic acid weight loss) was calculated from the weight and surface area. Lactic acid weight loss was calculated using Formula (1): Chemical composition analysis and observation of metallographic structures were performed using plate specimens prepared in the same manner as the preparation of the specimens for the immersion tests. Chemical composition analysis was performed by fluorescent X-ray analysis (XRF) and X-ray diffraction (XRD). XRF was performed with a fluorescent X-ray analyzer . XRD was performed using a desktop X-ray diffractometer . Surface structure was observed using a digital microscope . The observation was performed on three patterns: as sintered, after polishing, and after etching (acidic solution (nitric acid:hydrofluoric acid:water = 4 1:5) was used for etching).Each test was repeated six times from specimen preparation to measurement (n = 6). Two-way ANOVA was performed to analyze the results of the tensile test using DU ; factor B: building angle ). Three-way ANOVA was performed to analyze the results of the physical property tests using BL ; factor B: molding method ; factor C: surface condition (as sintered or after polishing)). Two-way ANOVA was performed to analyze the results of the immersion test ; factor B: molding method ). One-way ANOVA was performed on XRF for each detected element. From the ANOVA results, Tukey’s multiple comparison was performed when there was a significant difference for the measurement item.p < 0.01) only in the main effects . Subsequent Tukey’s multiple comparison test showed there was a highly significant difference (p < 0.01) in metal type (A) , but no significant difference (p > 0.05) in building angle (B). p < 0.05) than that of the SLS-0 specimen. The tensile strength of the specimens molded by SLS was significantly larger (p < 0.05) than that of the cast specimens at any building angle. For pure Ti, the tensile strength of the SLS-90 specimen was significantly larger (p < 0.01) than that of the SLS-0 specimen. Comparing the tensile strengths of the specimens molded by SLS with those of the cast specimens, the tensile strength of SLS-45 and SLS-90 specimens was significantly larger (p < 0.01) than that of the cast specimen.p < 0.01) only in main effects . Subsequent Tukey’s multiple comparison test revealed there was a highly significant difference (p < 0.01) in metal type (A) , but no significant difference (p > 0.05) in building angle (B). p > 0.05) in elongation with regards to the building angle of SLS specimens. The elongation of the specimens molded by SLS was significantly smaller (p < 0.05) than that of the cast specimen at all building angles. For pure Ti, there was no significant difference (p > 0.05) between the molding methods or among the building directions.Two-way ANOVA was performed on the elongation results, and revealed highly significant differences (p < 0.01) in the main effects B, C, and in the interaction B × C. p > 0.05).p < 0.01) in A, C, A × B, A × C, and A × B × C; and a significant difference (p < 0.05) in B. p <0.01) than that of the cast specimen. After polishing, these values were similar to those of the cast specimen before polishing. For pure Ti, the SLS-Z sintered contact angle was significantly smaller (p < 0.05) than that of the cast specimen. After polishing, this value was similar to that of the Ti-6Al-4V cast specimen before polishing.Regarding contact angle, there were highly significant differences (p < 0.05) in A × C, and a highly significant difference (p < 0.01) in all the other main effects and interactions. p < 0.01) than that of the cast specimen. After polishing, the Vickers hardness of the cast specimen was similar to those of the SLS-Z and SLS-XY specimens, and there was no significant difference between the molding methods or among the building directions (p > 0.05). The Vickers hardness of pure Ti was significantly smaller than that of Ti-6Al-4V in both SLS and casting method (p < 0.05).Regarding Vickers hardness, there was a significant difference (p < 0.01) in main effect A, but there was no significant difference (p > 0.05) in the molding method. The color difference of pure Ti was significantly increased (p < 0.01) compared with that of Ti-6Al-4V .Two-way ANOVA based on metal type (A) and molding method (B) was performed to assess their effect on the color difference (NBS unit) after immersion in either sodium sulfide or lactic acid solution, and lactic acid weight loss. Regarding the color difference after immersion in sodium sulfide solution, there was a highly significant difference (p < 0.01) in main effect A, but there was no significant difference (p > 0.05) in molding method. The lactic acid weight loss of Ti-6Al-4V was significantly (p < 0.01) larger than that of pure Ti .Regarding lactic acid weight loss, there was a highly significant difference (p < 0.01) in main effect A, and a significant difference (p < 0.05) in the interaction A × B. p < 0.05) larger than that of the cast specimen. For pure Ti, there was no significant difference between the molding methods or among the building directions (p > 0.05).Regarding the color difference after immersion in lactic acid solution, there was a highly significant difference (p < 0.05) higher than that of the cast specimen. The vanadium (V) fraction of the SLS-molded specimens was significantly (p < 0.05) lower than that of the cast specimen. The iron (Fe) fraction was in descending order of SLS-XY > SLS-Z > cast. The aluminum (Al) and chromium (Cr) fractions did not significantly differ (p > 0.05) between specimens of the different molding methods. For pure Ti, the Ti fraction was in descending order of SLS-Z > SLS-XY > cast. The Fe fraction of SLS-Z specimens was significantly lower (p < 0.01) than that of the cast specimen. For the zirconium (Zr) fraction, there was no significant difference between the molding methods or among the building directions (p > 0.05).In this study, several properties of the Ti-6Al-4V and pure Ti specimens produced using the SLS process were evaluated and compared to that of the casting specimens. Consequently, it was found that the specimens produced by SLS had comparable or superior properties compared with casting specimens. Besides, the building direction affected the tensile strength and surface roughness, but did not significantly affect other properties.SLS is considered a promising AM method for manufacturing metal dental prostheses. To date, Co-Cr alloy powder has been primarily used for SLS in dentistry. It is difficult to manufacture fine titanium powders, and the production process to produce AM-qualified raw material were technically complicated . For theThe tensile strength of Ti-6Al-4V was more than twice that of pure Ti. Moreover, the tensile strength values obtained for Ti-6Al-4V in this study satisfied those specified in JIS H4650 . For purThe elongation of pure Ti was larger than that of Ti-6Al-4V. The elongation values obtained in this study were close to those specified in JIS H4650   :cos θw =In this formula, θ is the contact angle, r is the area ratio of rough surface to plane (r ≧ 1), θw is the contact angle on the rough surface, and f is the ratio of liquid droplets entering the concave surface . Therefore, if f = 1, the larger the surface roughness, the larger the apparent contact angle on a hydrophobic surface (θ > 90°), and the smaller the apparent contact angle on a hydrophilic surface (θ < 90°). Therefore, for Ti-6Al-4V, the sintered contact angle of the SLS-molded specimen was large because its surface roughness was large. However, although the sintered surface roughness of the cast specimen was larger than that of the SLS-molded specimen, the contact angle of the cast specimen was smaller than that of the SLS-molded specimen. This could be because the f value is small; because the surface roughness of the cast specimen was larger; the concave surface could not be filled with water droplets. As a result, the contact angle of the cast specimen was small. For pure Ti, the surface roughness of the cast specimen was larger than that of the SLS-Z specimen. However, before polishing, the contact angle of the cast specimen was larger than that of the SLS-Z specimen. This could be because the effect of the f value was small. After polishing, the contact angles were 55°–70° for both metals. There was no difference between the molding method and metal type.For Ti-6Al-4V, before polishing, the Vickers hardness of the cast specimen (650.99 HV) was larger than that of the SLS-molded specimen. During casting, the metal reacted with the refractory and binder materials of the investment material to form a hardened layer containing impurities on the surface layer of the specimen. We consider that the Vickers hardness of the cast specimen was increased by this hardened layer. Polishing removed the hardened layer; therefore, after polishing, the Vickers hardness of the cast specimen was approximately 331–400 HV, which was hardness of the SLS-molded specimen before and after polishing. No significant difference was not observed between the molding methods or among the building directions. For pure Ti, before polishing, the Vickers hardness of the cast specimen (278.51 HV) was larger than that of the SLS-molded specimens. However, after polishing, it was 219.09 HV, which was not different from the values of the SLS-molded specimens before and after polishing.The results of the color difference after immersion in sodium sulfide solution did not differ between the molding methods. The value of Ti-6Al-4V was 12.64 (standard deviation: 5.04), which was slightly smaller than, but within the standard deviation of, that of 15.2 previously reported for Ti-6Al-4V . Further2; pure Ti: 56.68 µg/cm2). According to a previous report, when Ti-6Al-4V or pure Ti were immersed in lactic acid solution, the amount of Ti dissolved was 10 µg/cm2 and 23 µg/cm2, respectively [The molding method did not affect lactic acid weight loss. The value of Ti-6Al-4V was larger than that of pure Ti , there were no differences between the molding method for each alloy. The results of XRD were obtained using the plate-shaped specimens, not from the powder specimen, so the crystal orientation was not randomized. Therefore, the comparisons of the relative intensities of each peak were considered to be meaningless, so in this study, we focused only on the comparison on the positions of the peaks, namely the diffraction angles of the peaks (2θ). Consequently, it could be considered that the spectra of SLS specimens were not affected by the molding directions or molding methods in either Ti-6Al-4V or pure Ti. The diffraction angles of peaks obtained in this study were close to those previously reported in the literature ,33. For Ti-6Al-4V, there was no obvious difference in the surface structures of specimens made using the different molding methods. The pure Ti as sintered SLS specimen comprised a large block. In contrast, the cast specimen had a finer surface structure. However, these differences were limited to the surface layer. After polishing and etching, there were no differences in the surface structure between the molding methods.From the above, considering the mechanical, physical, chemical, and surface properties, Ti-6Al-4V and pure Ti specimens molded by SLS had either the same or better properties than cast specimens of Ti-6Al-4V and pure Ti. Only tensile strength and surface roughness were affected by the building direction of these specimens. Therefore, it can be concluded that the SLS process with either Ti-6Al-4V alloy powder or pure Ti powder is useful for clinical dentistry.The main limitation of this study is that the environment of the oral cavity was not considered. That should be more complex than the experimental environment, so the difference must affect these properties. Further study is needed to clarify them.In this study we compared the mechanical, physical, chemical, and surface properties of SLS-molded specimens using Ti-6Al-4V alloy powder and pure Ti powder with those of cast specimens of Ti-6Al-4V and pure Ti. The effect of the difference in the building direction of these specimens on the properties was also investigated. As a result, the following conclusions were obtained: (1) In both metals, the tensile strength of specimens with the same stress direction and building direction was small. The values of the specimens molded by SLS  were larger than those of the cast specimens  for both metals. (2) For Ti-6Al-4V, the elongation of the specimens molded by SLS (11.27%) was smaller than that of the cast specimen (14.72%). In pure Ti, both values were equivalent.(3) In both metals, the surface roughness of the surface of the building direction of SLS-molded specimens  was larger than other building direction . The value was smaller than that of the cast specimens . After polishing, there was no difference in surface roughness between the molding methods in both metals.(4) For Ti-6Al-4V, the sintered contact angles of the SLS-molded specimens (98.14°) were larger than those of the cast specimen (68.86°). In Ti-6Al-4V after polishing and pure Ti, there were no obvious differences in the molding method.(5) For Ti-6Al-4V, the sintered Vickers hardness of the SLS-molded specimens (366.84) was less than that of the cast specimen (650.99). In Ti-6Al-4V after polishing and pure Ti, there were no differences in the molding method.(6) In both metals, there were no differences between the molding methods regarding color difference after sodium sulfide immersion. The color difference of pure Ti was larger than that of Ti-6Al-4V.(7) In both metals, there were no differences between the molding methods regarding lactic acid weight loss. Lactic acid weight loss in Ti-6Al-4V was larger than in pure Ti.(8) For Ti-6Al-4V, the color difference of the SLS-molded specimens after lactic acid immersion was greater than that of the cast specimen. In pure Ti, the values were small, and there was no difference between the molding methods.(9) In both metals, there were no obvious differences between the molding methods for XRF, XRD, and surface structure.From the above, it is concluded that SLS molding using either Ti-6Al-4V alloy powder or pure Ti powder is an extremely excellent molding method for clinical dentistry.


DSM-5 using a 5-point Likert scale. This study investigated its validity and reliability in screening for BPD in university students.The screening instrument for borderline personality disorder (SI-Bord) consists of a 5-item self-reported questionnaire on the key features of BPD from the A cross-sectional study was conducted on a sample of university students in Thailand between November and December 2019. An online assessment gathered demographic data and results from the SI-Bord, the Perceived Stress Scale-10 (PSS-10) and the Patient Health Questionnaire-9 (PHQ-9). Participants whose SI-Bord scores were ≥ 1 were randomly selected to be interviewed and assessed for a BPD diagnosis by four psychiatrists using the Structured Clinical Interview for DSM-IV Axis II Personality Disorders (SCID-II) as a reference point. An intraclass correlation coefficient (ICC) of 0.925  ensured inter-rater reliability between the four psychiatrists. The diagnostic sensitivity and specificity of the SI-Bord, as compared to that of the SCID-II, were determined to indicate the cut-off score. The Receiver Operating Characteristics (ROC) was analyzed to evaluate its diagnostic accuracy.r = 0.67 and r = 0.69, p < 0.001, respectively) and negatively correlated to MSPSS . The prevalence of BPD in the sample was 6.4%, according to the cut-off score > 9.The study included 342 students aged 18–25 years (the mean age was 20.25 ± 1.4 years), 80.4% of whom were female. Among the 68 participants selected for an online interview, 16 were diagnosed with BPD. The cut-off score of the SI-Bord was > 9, as suggested by the Youden index, yielding a sensitivity of 56.3% and a specificity of 92.3%. It had a positive predictive value of 69.2% and negative predictive value of 87.3%. The SI-Bord had adequate discriminative power between cases and non-cases of BPD, with the area under the ROC curve being 0.83. Cronbach’s alpha for the SI-Bord was 0.76, indicating acceptable internal consistency. The SI-Bord score was positively correlated to PHQ-9 and PSS-10 scores (The SI-Bord demonstrated good reliability and validity for screening BPD in university students. However, a study in non-Thai and other population groups should be warranted. As a group, university students commonly experience stress and depression as they transition from adolescence to early adulthood, which often requires them to adapt to a new social role and identity, to maintain interpersonal relationships, to manage their own finances, and to strive for academic success . People Diagnostic and Statistical Manual of Mental Disorders, fifth edition (DSM-5), BPD is characterized by ‘a pervasive pattern of instability of interpersonal relationships, self-image, and affects, and marked impulsivity beginning by early adulthood and present in a variety of contexts’ [One important factor associated with suicidality is borderline personality disorder (BPD), which usually become apparent during adolescence and young adulthood , 9. Accoontexts’ . BPD is ontexts’ –13 and tontexts’ . BPD is ontexts’ .As for the prevalence of BPD among university students, a review of 43 studies reported that the prevalence of BPD ranged from 0.5 to 32.1%, with an unadjusted lifetime prevalence of 9.7% . It is, Up to 13 measurements are used to diagnose BPD  in widelDSM-5 criteria. Short-Bord was tested in psychiatric patients who came for psychotherapy and showed an AUC of 0.95, a sensitivity of 91.2% and a specificity of 85.4%, determined by a cut-off of ≥2, and a Cronbach’s alpha of 0.80 [Recently, the authors  developed a short screening tool for borderline personality disorder, comparable to the BPD subscale in SCID-II (Short-Bord) that uses a true-false questionnaire with five items based on  of 0.80 . AlthougThis version – the short instrument for borderline personality disorder (SI-Bord) – is, therefore, an improved version of Short-Bord, which was reliable and valid. Compared with other current screening instruments, the SI-Bord is shorter and culturally validated . The objectives of this study were to administer the SI-Bord to a sample of university students to 1) test the diagnostic validity of the SI-Bord, and 2) test its reliability and validity compared to other psychological measurements.This study used a cross-sectional online survey of 342 undergraduate students in Thailand between November and December 2019. To participate in the online survey, the participant had to read the protocol and accept an informed consent document on the first page of the questionnaire. Participants who objected to the informed consent document were directed to the end of the survey. No respondent was forced to participate and he/she could withdraw at any time. For the interview and video recording of the interview, both the interviewer and the participant were informed that the proceedings would be private and confidential. The study was approved by the Ethics Committee of the Faculty of Medicine at Chiang Mai University, Thailand.The participants were undergraduate students located all over Thailand, aged18–25, who spoke Thai. The exclusion criteria included being diagnosed with schizophrenia, bipolar disorder, drug or alcohol use disorder, and being intoxicated with alcohol within 24 h prior to participating in the study. Participants were asked to complete the questionnaires on the Internet via personal computers, laptops, smartphones, or tablets.Those who scored ≥1 on the Short-Bord were randomly selected to participate in a long-distance interview (using Zoom video conferencing) by one of the psychiatrist investigators  to confirm the diagnosis using the Structured Clinical Interview for DSM-IV Axis II PDs (SCID II). Video of the interviews was recorded for later review.Sample size estimation was calculated for a prevalence study, which the estimated prevalence of BPD in the university sample from previous study was 30%, the precision was set at 5%, and the confidence level was 0.95. This yielded an estimated sample size of 323. The dropout rate was determined to be 10%. The estimated total sample was 355 for the survey.To calculate a sample size for ROC analysis, we expected an AUC of 0.725 for the SI-Bord, which was significant compared to the null hypothesis value 0.5 (no discriminating power). We expected to include thrice as many negative cases as positive cases based on the prevalence. An α-level was set to 0.05, and a β-level was set to 0.20 (power is 80%). This yielded a sample size of 68  .  in the process of interrater reliability. All ten cases were interviewed by four psychiatrists. Two psychiatrists interviewed two cases each, and three psychiatrists interviewed three cases each. Each case was interviewed by one interviewer, and the other three psychiatrists rated the symptoms independently by watching the videos. All four psychiatrists were blinded to all cases. This totaled six pairs of interviewers for each video, and 60 pairs for all comparisons. No SCID-II Personality Questionnaire was used before any interview. The intraclass correlation analysis yielded a coefficient of 0.925, which was considered excellent.Screening instrument for borderline personality disorder (SI-Bord)In addition to the sociodemographic data, e.g., age, sex, academic year, income, etc., the participants were asked to complete the following measurements.2)Revised Thai Multi-Dimensional Scales of Perceived Social Support (r-MSPSS)SI-Bord was modified from the Short-Bord , consist3)Thai version of Perceived Stress Scales (T-PSS-10)This tool measures the extent to which an individual has experienced being support by significant others (SO), friends (FR), and family (FA) . It has 4)Patient-Health Questionaire-9 (PHQ-9)The T-PSS-10 is a 10-item self-reporting questionnaire measuring the extent to which an individual perceived stress over the past 4 weeks. It uses a 5-point Likert scale ranging from never (0) to very often (4). The higher the total score, the higher the level is attained of feeling stress. T-PSS-10 demonstrates good psychometric properties . The stu5)Thai version of Structured Clinical Interview for DSM-IV Axis II PDs (T-SCID II)The PHQ-9 is a 9-item self-reporting questionnaire measuring the extent to which an individual has experienced depressive symptoms over the past 2 weeks . The 4-rT-SCID-II is a structured clinical interview for personality disorder that has been used to diagnose personality disorders based on DSM-IV and DSM-IV TR . The ThaJ [c + specificityc - 1), where c ranges over all possible criterion values. MedCalc Statistical Software, Version 19.2.1  was used for ROC analysis. MedCalc was used to calculate the optimal criterion and associated sensitivity and specificity, and the optimal criterion value which takes into account not only sensitivity and specificity, but also disease prevalence, and costs of various decisions. Cost, as defined by MedCalc, is the average cost resulting from the use of the diagnostic test at that decision level such as the cost of doing the test, which is constant at all decision levels. ROC analyses were also used for the area under the curve calculation (AUC) to determine the performance of the test.For sociodemographics and scores of psychological measurement, descriptive statistics, e.g., frequency, percentage, mean and SD were used. In evaluating the diagnostic validity of the SI-Bord, receiver-operating characteristics (ROC) analyses were used to calculate the values of sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV). We calculated the Youden index J  which isInterrater reliability calculated by intra-class correlation coefficient (ICC) was used for rater’s agreement. An ICC value of > 0.8 was acceptable. Internal consistency of the instrument was determined by Cronbach’s alpha, for which a value of > 0.7 was considered acceptable. Concurrent validity of the SI-Bord was examined using Pearson’s correlation coefficients between SI-Bord and with other measurements, i.e., rMSPSS, T-PSS-10, and PHQ-9. All analyses, except for ROC, were performed using IBM SPSS, Version 22.The total number of responses was 355. Among them, 13 were excluded: five respondents were older than the inclusion criteria (25 years old) and eight were repeat responses. The final number of participants was 342, with 275 females (80.4%) and a mean age of 20.25 (SD = 1.4). The majority of the participants were health science students. Among the 68 participants who were interviewed, 45 were females and 23 were males. Sixteen  participants met the criteria for BPD. Other characteristics are shown in Table p < 0.0001), suggesting an 83% chance that the SI-Bord will correctly distinguish a BPD case from a non-BPD case. Table The ROC curve was analyzed to determine the cut-off score of the SI-Bord Fig. . The AUCr = 0.598, p < 0.001), while item 2 (unstable relationship) had a nonsignificant correlation with the psychiatrists’ diagnoses . Factor analysis results show that a one-factor solution model fits the data best. The total variance explained by all items was 51.42% with an eigenvalue of 2.57. The factor loading coefficient ranged from 0.548 (item 1) to 0.832 (item 5), consistent with the strength of the item-total correlation coefficient, which ranged from 0.376 (item 1) to 0.660 (item 5).Table r = 0.669 to r = 0.690, p < 0.001) but was negatively correlated with perceived social support .Table The aim of this study was to test the efficacy of the SI-Bord on university students, and the results showed it is both valid and reliable for this population. Most importantly, the results indicate that the SI-Bord is satisfactory in its diagnostic accuracy, which is comparable to other vetted screening questionnaires such as the BPQ (AUC = 0.83) and the BDQ (AUC = 0.85) , despiteThe SI-Bord’s AUC of 0.83 means that a randomly selected individual from the positive group will have a test value larger than that of a randomly chosen individual from the negative group 83% of the time . CompareDSM criteria. As a screening tool, only those items that correlate strongly with the reference diagnosis were counted [Unlike MSI or IPDE, the SI-Bord uses only the most important diagnostic criteria of BPD instead of using all 9  counted , 41. It  counted , 43. Nee counted , 45, ren counted .Despite its unidimensional scale, item 2 (unstable relationship) on the SI-Bord was not productive for the scale as it could not discriminate between cases and non-cases. It had a nonsignificant correlation with the psychiatrists’ diagnoses despite being uniformly consistent with the other four items as indicated in previous research . This reAs a screening tool, the SI-Bord gives clinicians the option of using various cut-off scores depending on the clinical purpose. For example, to increase the probability of including a case, a cut-off score of > 7 has higher sensitivity (75.0%) than the cut-off of > 9 (56.25%) but using the lower cut-off score could increase the chance of a false positive. The negative predictive value differed little between the two scores (90.5% vs 87.3%) but the positive predictive value differed greatly (46.2% vs 69.2%). Therefore, a cut-off of > 9 would be preferred over the cut-off > 7 when researchers are interested in excluding non-cases from the study . In addiOur study indicates that the validity of the SI-Bord is similar to other psychological measurements and one interpretation of our results is that a high score on the SI-Bord, is, as expected, associated with a higher level of depression and perceived stress and a lower level of perceived social support, which is consistent with other related studies , 50. ThiThe SI-Bord is, to the best of our knowledge, the first screening tool for BPD specifically designed to screen university students. The SI-Bord is a unidimensional scale that, although short, is comparable with longer scales in terms of its psychometric properties and its ability to distinguish between participants with and without BPD.This study has some limitations. The information obtained in the interviews using the SCID-II diagnosis was limited, as it was obtained only from the participants and only used participants’ self-reported history to exclude those with clinical disorders . The accuracy of the data may have been impacted by a lack of corroborating data. Furthermore, the data in this study were obtained from participants with access to computers and other smart devices. Those who were excluded because they were unable to access the survey may also be at risk for BPD. This study did not exclude other common comorbidity in Asia such as Internet addiction that was found to be common in university students  and adolThe SI-Bord showed good validity and reliability in screening for BPD. It is a brief and reliable screening tool for university students. Further investigation should be encouraged, especially to determine which items yield the most accurate and reliable results.Additional file 1: Appendix. Screening instrument for borderline personality disorder (SI-Bord).


Between baseline and 4 weeks Total Cholesterol (TC) levels showed an average reduction of 6.5% (95%CI: −10.9 to −1.9) in the beta-glucan sequence; while non-HDL-C plasma concentrations decreased by 11.8% (95%CI: −14.6 to −4.5). Moreover, after 8 weeks of beta-glucan supplementation TC was reduced by 8.9% (95%CI: −12.6 to −2.3) and non-HDL-C levels by 12.1% (95%CI: −15.6 to −5.3). Decreses in TC and non HDL-C were significant also versus placebo (respectively p < 0.05 and p < 0.01 to both follow-up visits). Fasting plasma glucose and self-perceived intestinal well-being were not affected by both beta-glucan and placebo supplementation.The Beta-glucan Effects on Lipid profile, glycemia and inTestinal health (BELT) Study investigated the effect of 3 g/day oat beta-glucans on plasma lipids, fasting glucose and self-perceived intestinal well-being. The Study was an 8-week, double-blind, placebo-controlled, cross-over randomized clinical trial, enrolling a sample of 83 Italian free-living subjects, adherent to Mediterranean diet, with a moderate hypercholesterolemia and a low cardiovascular risk profile. Beta-glucans reduced mean LDL-Cholesterol (LDL-C) levels from baseline by 12.2% (95%CI: −15.4 to −3.8) after 4 weeks of supplementation and by 15.1% (95%CI: −17.8 to −5.9) after 8 weeks of supplementation ( The consumption of dietary supplements to control plasma cholesterol levels has become widespread in recent years, alongside the publication of several systematic reviews and meta-analyses showing the cholesterol lowering effect of some of these products, including fibers ,2,3.A comprehensive meta-analysis of 58 clinical trials and 3974 subjects has recently showed that oat beta-glucan significantly affects the serum concentrations of low-density lipoprotein cholesterol (LDL-C), non-high-density lipoprotein cholesterol (non-HDL-C) and apolipoprotein-B (apo-B), concluding that the inclusion of oat-containing foods in the diet may be a valid strategy to prevent the onset of cardiovascular disease . In partThe mechanisms underlying the lipid-lowering properties of dietary fiber are still not fully understood . The abiThe contribution of beta-glucans from oat and barley to the maintenance of normal blood cholesterol levels and their efficacy in the reduction of blood cholesterol levels at a dosage of 3 g per day was formally recognized by the European Food Safety Authority (EFSA) following specific applications and health claims that were authorized in the European Union (Commission Regulation (EU) 432/2012) . HoweverIn this context, we deemed interesting to evaluate the effect of a proprietary formulation of beta-glucans, at the dosage of 3 g/day on fasting plasma lipids and glucose, as well as its tolerability, in an Italian sample of subjects with mild hypercholesterolemia.The Beta-glucan Effects on Lipid profile, glycaemia and inTestinal health (BELT) Study was a medium-term, double-blind, placebo-controlled, cross-over randomized clinical trial, which enrolled a sample of Italian free-living subjects with moderate hypercholesterolemia recruited from the Lipid clinic of the S. Orsola-Malpighi University Hospital, Bologna, Italy.2), liver failure, renal failure (estimated glomerular filtration rate (eGFR) < 0.5 mL/s), irritable bowel syndrome, inflammatory bowel diseases and food allergies, as well as having been treated in the previous 2 months with fiber based dietary supplements and/or probiotics and/or lipid-lowering drugs or any other drugs potentially able to affect the lipid metabolism.Participants were required to be aged between 20 and 65 years, with moderately high levels of TC (TC ≥ 5.17 mmol/L and ≤ 6.2 mmol/L) and LDL-C (LDL-C ≥ 3.36 mmol/L and ≤ 4.91 mmol/L) and an estimated 10-year cardiovascular risk <10%, as per the country-specific risk charts from the CUORE project [Participants were adhering to a standardized diet for four weeks before being randomized to receive adequate supplies of either the beta-glucan supplement or placebo, in order to complete the one of two 2-month treatment sequences. The crossover to the second treatment was preceded by a 4-week wash-out period. Finally, participants were asked to return for follow-up visits two and four weeks after stopping supplementation. The study timeline is described in detail in At enrollment, subjects were instructed to follow the general indications of a Mediterranean diet, avoiding an excessive intake of diary and red meat derived products.Once every two visits, subjects were provided with a food diary to record their 3-day intake (two week-days and one week-end day), which they were requested to complete and return during their subsequent visit. In particular, the food diaries were collected and the diets analyzed at the beginning and at the end of each treatment phase.®) based on a large food database that is frequently updated with values from the main official Italian databases (INRAN/IEO 2008 revision/ADI). Data were handled in compliance with the company procedure IOA87.The analysis of diet composition was performed using a dedicated software .The study was conducted in accordance with the Good Clinical Practices and fully complied with the ethical guidelines of the Declaration of Helsinki. All subjects received an informational document describing the study and signed a consent form for study participation. The study was approved by the Bologna University Ethical Committee (Code: BELT_2016) and registered on ®, Herbalife) or an oat-based isocaloric placebo without beta-glucan. The tested products had a similar macronutrient composition  and were indistinguishable in color and taste. In particular, the Oatwell™ composition per 100 gr included 22 gr of beta-glucan soluble fiber characterized by very high molecular weight polysaccharides (>2000 kDa) and high viscosity  , physica2).Waist circumference (WC) was measured at the end of a normal expiration, in a horizontal plane at the midpoint between the inferior margin of the last rib and the superior iliac crest. Height and weight were measured to the nearest 0.1 cm and 0.1 kg, respectively, with subjects standing erect with eyes directed straight wearing light clothes and with bare feet. BMI was calculated as body weight in kilograms, divided by height squared in meters  readings were sequentially obtained at 1-min intervals. The first one was then discarded, and the average between the second and the third was recorded .2EDTA) (1 mg/mL) and blood centrifugation at 3000 RPM for 15 min at room temperature.The biochemical analyses were carried out on venous blood, withdrawn after overnight fasting (at least 12 h). Plasma was obtained by addition of disodium ethylenediaminetetraacetate  version 22.0  for Windows.A full descriptive analysis of the collected parameters was carried out as per protocol. Categorical variables were expressed as absolute number and percentage and compared with the Fisher corrected chi-square test or the Wilcoxon-Rank test, based on whether they were nominal or ordinal. Continuous variables were expressed as mean ± standard deviation (SD) or mean and standard error (SEM), and compared by analysis of variance (ANOVA) followed by post-hoc Tukey test or by Kruskal-Wallis non parametric analysis of variance followed by Dunn’s pairwise test, depending on their statistical distribution . To verify the basic assumption of cross-over design, the presence of a carryover effect was excluded.p < 0.05 two-tailed. The Dixon’s Q test was always carried out to exclude the extreme values.The minimum level of statistical significance was set to Efficacy analyses were performed considering the intention-to-treat (ITT) population, i.e., all subjects with at least one post-baseline control. A sensitivity analysis of the primary variable was also planned in the per-protocol population, i.e., all subjects without major protocol violations.2 vs. ≥ 27 kg/m2).The primay efficacy outcome analysis was also repeated by gender, age group (20–40 vs. 40–65 years old) and weight group . Furthermore, the study groups were well matched for all the considered variables at baseline.All enrolled subjects  successfully completed the trial according to the study design. The final distribution between men and women, when considering the treatment sequences, did not show any significant differences , which corresponded to an absolute decrease of 0.59 mmol/L (95%CI: −0.8 to −0.39) at the end of the intervention period (p = 0.04), in younger subjects (16.4% (95%CI: −17.5 to −8.3) vs. older 14.7% (95%CI: −17.1 to −5.2)) while no difference has been detected as regards body mass index classes (p > 0.05).On the contrary, the active supplementation reduced mean LDL-C levels from baseline by 12.2% (95%CI: −15.4 to −3.8) after 4 weeks and by 15.1% (95%CI: −17.8 to −5.9) after 8 weeks  in the beta-glucan sequence, while non-HDL-C plasma concentrations decreased by 11.8% (95%CI: −14.6 to −4.5). Moreover, after 8 weeks of treatment, beta-glucan reduced TC by 8.9% (95%CI: −12.6 to −2.3), corresponding to an absolute decrease of 0.52 mmol/L (95%CI: −0.72 to −0.32), and non-HDL-C levels were reduced by 12.1% (95%CI: −15.6 to −5.3), which corresponded to 0.53 mmol/L (95%CI: −0.73 to −0.33) .Descriptions and results of exploratory analyses of the other secondary endpoints, the effect on which were considered non-significant, are provided in After treatment discontinuation, both TC and LDL-C rapidly reversed to basal values. Lipid plasma levels assessed after 2 weeks of washout were comparable with those measured at baseline and not significantly different from those observed at the end of the wash-out period .The compliance with the treatment was almost complete (89%) during both treatment periods.The tolerability profile of the beta-glucan supplement used was rated as acceptable by most subjects. During supplementation with beta-glucan, three subjects experienced moderate abdominal discomfort  and one subjects experienced dysphagia. No one experienced adverse events regarding laboratory parameters during the trial. No volunteer discontinued the trial because of adverse events that occurred during the treatment and no effects reported in conjunction with use of the placebo.In particular, the tested products did not exert any significant unvafourable effect on the self-perceived intestinal well-being. The results of the second-ranked secondary endpoint are summarized in Beta-glucans  are polysaccharides naturally occurring in the cell wall of grains and cereals, especially barley and oats . In our The observed effect on TC and LDL-C are larger  than expected, based on the most recent meta-analysis and EFSA opinion, which estimate respectively a mean change in LDL-C of 0.3 mmol/L and 0.21 mmol/L (about 7–10% of baseline concentratios) . The reaFurthermore, the observed effect might also be due to the characteristics of the considered population sample. In this regard, a reliable meta-regression analysis recently demonstrated a significant inverse association between baseline LDL-C levels and the extent of LDL-C reduction after supplementation with oat beta-glucan . These rSimilarly, the lack of any significant effect on FPG in the trial might be due to the characteristics of the enrolled subjects, being all euglycemic. Actually, a recent meta-analysis found that oat beta-glucan intake was more effective in people with type 2 diabetes .a priori more attentive to a healthy lifestyle) and to the run-in stabilization diet period preceding the treatment phase, during which possible dietary mistakes were corrected before the start of the trial. Thus, the results obtained with beta-glucan supplementation are more representative of what would be observed in a setting of clinical practice.The lack of changes in metabolic parameters during supplementation with placebo may be attributed to both the enrollment in the setting of a lipid clinic (and consequently, presumably, of subject At the same time, the lack of effects of beta-glucans on intestinal function parameters in our study should be related to the Mediterranean diet pattern of the enrolled subjects and the exclusion from the enrolment of subjects affected by irritable bowel syndrome, inflammatory bowel diseases and food allergies, but also because of the low amount of supplemented fibers.The observed results are largely supported by the mechanisms of action by wich beta-glucans can improve cholesterolemia. The LDL-C lowering effect of beta-glucans has mainly related to the their ability to act as dietary fibers, consequently entrapping bile acid micelles, impairing their ability to interact with luminal membrane transporters on the intestinal epitelium, thereby increasing the fecal cholesterol output.  The follThe BELT Study has some relevant limitations. For instance, the observational period was not long enough to assess any adaptation phenomena possibly occurring by changes in the gut microbiota composition. For this reason, further longer-term clinical studies are needed to confirm if the dietary intake of beta-glucan enriched foods is able to maintain the observed positive metabolic effect over the time.Secondly, we did not evaluate any marker of intestinal absorption during the study, since the aim of this study was a clinical and not a pharmacological evaluation of the effects of supplementation with beta-glucan-enriched dietary supplement in moderately hypercholesterolemic subjects.However, the BELT Study is the first placebo-controlled clinical trial testing the effect of oat beta-glucan supplementation on a large sample of a South European population strictly adherent to the Mediterranean diet. This is of particular interest because the high fiber content of the Mediterranean diet might have theoretically reduced the effect of beta-glucan supplementation on serum lipids , while tIn conclusion, the BELT study confirms the medium-term efficacy of supplementation with 3 g/day of beta-glucan in reducing LDL-C, TC and non-HDL-C in mild hypercholesterolemic subjects, even in the context of a Mediterranean setting.


Temporal estimation can be influenced by pain, which is a complex psychological and physiological phenomenon. However, the time range in which perception is most sensitive to pain remains unclear.In the present study, we explored the effects of acute inflammatory pain on time perception in the sub- to supra-second (0.6–2.4-s) and supra-second (2–8-s) ranges in rats. Plantar formalin injection was used to induce acute inflammatory pain, and a temporal bisection task was used to measure time perception. Task test sessions were held for five consecutive days (one per day): the day before injection (baseline), immediately after injection, and the three post-injection days. The point of subjective equality  and Weber fraction  were calculated and analysed.p = 0.049) immediately after injection. Formalin-induced pain also tended to lengthened time perception in the 0.6–2.4-s range on post-injection days 2 (p = 0.06) and 3 (p = 0.054). In the 2–8-s range, formalin injection did not affect the PSE or Weber fraction.In the 0.6–2.4-s range, the PSE was significantly lower, indicating prolonged subjective duration, in the formalin group relative to the saline group (The enhanced effect of pain on temporal perception in the sub- to supra-second range is observed in this study and this effect is attenuated with the prolongation of estimated time, even in rats. Pain plays an important role in protecting individuals from potential or further injury . Stress Considerable number of evidences suggest that the perception of short but not long duration is modulated by negative emotional stimulus; the greatest effect has been observed for short-term (2-s) time perception, with a gradual decrease with the prolongation of duration (to 4–6 s) . The reaAs a complex physiological and psychological phenomenon with sensory, emotional and cognitive components , pain haIn human studies, long temporal ranges are affected easily by strategies such as counting . In ordeThirty-two male Sprague–Dawley rats  were used in this study. The rats were allowed to adapt to the laboratory environment for at least 1 week before the experiment was conducted, and they were handled (by stroking them or letting them move freely on the experimenter’s arms for 5 min) once per day to familiarise them with manipulation by the experimenter. They were housed individually in separate cages with food and water available ad libitum; but since the rats were able to get water from the task, we removed the water bottle from the feeding cage the day before the formal experiment. When a rat did not perform the task on any day, it was allowed to drink freely from a standard 250-ml bottle for 5 min. In addition, we recorded the rats’ body weights every 3–4 days and gave them an additional 5 min access to water when a loss of >10 g from the last recording was noted. During the experimental period, the rats’ body weights were maintained between 270 and 360 g.The room in which the animals were housed was temperature and humidity controlled , with a 12/12-h light/dark cycle (lights on at 8:00 pm). All experiments were performed in accordance with the National Institutes of Health guidelines for the care and use of laboratory animals  and approved by the Institutional Review Board of the Institute of Psychology, Chinese Academy of Sciences. The approve number is H16036. All rats were alive upon completion of this research and were used in preliminary experiments for another study.Temporal discrimination training and temporal bisection testing of each rat were conducted in the same operant box (21 cm W × 32.5 cm L × 42.5 cm H) located in a soundproof chamber. The box was made of acrylic, had a strip floor and was positioned above a waste catch pan. Two retractable levers  were mounted symmetrically on one wall of the box, 9 cm above the floor. A liquid dispenser (ENV-201A) and water receptacle (2.5 cm above the floor) were spaced equally between the levers to provide water as the reinforcer. As the to-be-timed auditory cue, a pure-tone stimulus  was presented by a tone generator (ENV-223AM) mounted on the wall above the liquid dispenser. An illuminated infrared detective nose-poke hole (ENV-114BM) was mounted in the middle of the wall opposite the levers, 2.5 cm above the floor. An indicator lamp (ENV-221M) was located directly above the nose-poke hole. An exhaust fan was mounted on the soundproof chamber. The operant box was connected to a computer that recorded the rats’ experimental events. The experimenter furnished all output and input devices mounted in the box. The box was cleaned with water and medical-grade alcohol after each session.n = 8 each) according to their baseline performance. All experiments were performed during the rats’ dark cycle.The temporal bisection task was conducted with 16 rats each for the sub- to supra-second and supra-second ranges. After training, each rat was first submitted to three test sessions (one per day) to stabilise its performance. Then, each rat performed five test sessions on five consecutive days (one per day): the day before formalin injection (baseline), immediately after formalin injection, and 1–3 days after formalin injection. Before the day of injection, the rats were assigned to formalin and saline control groups , each tone was triggered by a rat’s nose-poke behaviour. Only the corresponding lever was extended following the termination of the tone. For the free-choice response training trials (n = 100), the tones were triggered by the rats’ nose-poke behaviours, and their termination was followed by the simultaneous extension of both levers. On the first 6 days of free-choice training, the levers were retracted only after a rat provided the correct response; on the next 6 days, the levers were retracted immediately after any response. Correct responses were followed by the provision of drips of water as a reinforcer. Non-response for 5 s resulted in lever retraction and no reward. The criterion for the successful completion of temporal discrimination training was the achievement of correct response rates ≥ 85% on 3 consecutive days. For all training and testing sessions, the rats were allowed to move freely in the operant box.Temporal discrimination training was performed using a procedure modified according to previous research . It consDuring the testing stage of the temporal bisection task, a series of to-be-timed tones of anchor and intermediate durations was presented, and the rats were free to press any of the levers. In the testing sessions, each trial began with the illumination of the indicator light. The to-be-timed tone was initiated by a nose-poke. At the end of the tone, the two levers were extended simultaneously. Lever retraction was prompted by any lever press or non-response for 5 s. The average intertrial intervals  per rat and session were 20 s  for the 0.6–2.4-s range and 17 s  for the 2–8-s range. One testing session consisted of 140 trials : 14 trials each with tones of five intermediate durations without reinforcement, and 35 trials each with the two anchor-duration tones with reinforcement. Typically, the rats obtained about 65 reinforcements per testing session. For the sub- to supra-second–range (0.6–2.4-s) experiment, the five intermediate durations were set at 0.756, 0.952, 1.2, 1.51 and 1.9 s. For the supra-second range (2–8-s) experiment, the intermediate durations were set at 2.51, 3.17, 4, 5.04 and 6.35 s. The sequence of these trials was random, but with the constraint of no more than three consecutive intermediate-duration trials.Acute inflammatory pain was induced by injection of 50 µl 1% formalin solution into the plantar surface of each rat’s hind paw. An equal-volume saline injection was used as the control. After injection, the rats were returned to the operant box to perform the bisection task, and their nociceptive behaviours (paw licking and paw lifting) during the 70-min test sessions were video recorded. The durations of paw lifting and licking, measured by the experimenter in 5-min epochs, and the cumulative duration of nociceptive behaviours in phases I (0–10 min) and II (20–60 min) were calculated for analysis.PL,calculated by dividing the number of responses to the “long” lever by the total number of responses in trials with the same tone duration) to each to-be-timed tone in each temporal bisection session was used to analyse time perception. Curve fitting and parameter calculation were performed using the Prism software . Following previous work (F (t) is the PL when the duration is equal to a given sample (t), µ  is the mean, and σ is the standard deviation (representing the slope of the function). Parameters a represents the low asymptote and b represents the range of function. The mean (µ) is also the PSE, defined as a 50% chance that the animal will provide a “long” response (PL = 50%). The PSE reflects the subjectively perceived length of time, with increases and decreases therein interpreted as under- and overestimation, respectively, of the duration  or (day * duration)]; PSE, Weber fraction and omitted trials at baseline and on the injection day (group * day). Independent-sample t test was used to compare the differences of the cumulative duration of paw licking and lifting, the daily correct response rates, and the omitted trials and the changes in PSE/Weber fraction on each post-injection day between the two groups. If these data do not conform to the assumptions of the statistical method, we will use the correction method. The data are expressed as means ± standard errors of the mean, and the significance level was set at α <0.05. Data from trials with no lever-press response were omitted from the analysis.Changes in the Weber fraction in the post-injection sessions were calculated using the same formula. The Prism  and SPSS  software packages were used for graph generation and statistical analyses, respectively. Two-way repeated measure analysis of variance (two-way RM ANOVA) followed by a simple main-effect analysis was conducted to analyse the following variables: nociceptive behaviours (group * time); F  = 8.624, p < 0.001; treatment effect: F  = 28.500, p < 0.001; time effect: F  = 10.844, p < 0.001; t (14) = −5.711, p < 0.001; t (14) = −4.433, p < 0.001; As shown in t tests revealed no difference in the correct response rates at baseline or on the injection day, indicating that formalin treatment did not affect the animals’ temporal discrimination ability.We compared the correct response rates to anchor-duration tones between two groups at baseline  and on the injection day , respectively. Independent-sample PLs of temporal bisection task was presented in PLs at baseline  and a simple main-effect analysis showed that PLs on the injection day were higher than these at baseline at 0.952 s (p = 0.030) and 1.2 s (p = 0.017). No such effect was observed in the saline group . R2 values for curve fitting ranged from 0.86 to 0.99 (average: 0.94) in the two groups at baseline and on the injection day, with no difference between groups.The effect of formalin injection on F  = 6.405, p = 0.024]. A simple main-effect analysis showed that the PSE was lower in the formalin group than in the saline group on the injection day , but not at baseline . The Weber fraction, however, did not change significantly . The response latency (defined as the time between lever extension and pressing) did not differ between groups at baseline . The cumulative duration of paw licking and paw lifting was significantly greater in the formalin group than in the saline group in phases I  or on the injection day . However, the treatment had a significant main effect on the injection-day PL; no such effect was observed at baseline . The significant duration effect revealed that the rats discriminated the durations of the to-be-timed tones (p < 0.001). For each group, two-way RM ANOVA was also used to compare the differences of PLs between baseline day  = 0.804, p = 0.573] or saline group . R2 values for curving fitting ranged from 0.81 to 0.99 (average: 0.95) in the two groups at baseline and on the injection day, with no difference between groups [data from one rat were excluded because the R2 values from all tests were low (0.734 ± 0.168)].The effect of formalin injection on line day  and injeline day . No signF  = 0.772, p = 0.396] or the Weber fraction . At baseline, response latency did not differ between groups ; no such interaction was observed at baseline . The main treatment effect also was not significant . The main effect of duration was significant in all tests , indicating that the rats could discriminate the durations of the to-be-timed auditory stimuli. R2 values for curve fitting ranged from 0.87 to 1 (average: 0.96), with no significant difference between groups on any of the 3 days.The results of temporal bisection tasks performed in the 0.6–2.4-s range on post-injection days are presented in t tests showed that the formalin treatment tended to reduce the change of PSE on post-injection days 2  and 3 , see As shown in t tests revealed no post-effect of formalin treatment in the omitted trials from any day. Two-way RM ANOVA (duration * treatment) revealed that the treatment had no significant effect on response latency on any of the 3 days . The effect of treatment on PL also was not significant . Duration had a significant effect on the PL in all tests (p <0.001). R2 values for curve fitting ranged from 0.83 to 1 , with no significant difference between groups on any of the 3 days.The results of temporal bisection tasks performed in the 2–8-s range on post-injection days are shown in As shown in t tests revealed no effect of the formalin treatment on omitted trials from any day. The response latency did not differ significantly on post-injection days 1 and 2 [treatment: F  ≤ 2.58, p ≥ 0.132; interaction of duration and treatment: F  ≤ 1.365, p ≥ 0.239; F  = 0.913, p = 0.357; interaction of duration and treatment: F  = 2.666, p = 0.021; p = 0.022).Omitted trials from the three post-injection days  were analysed. Independent-sample In this study, we investigated the effect of formalin-induced pain on time estimation at two scales in rats. Plantar formalin injection lengthened subjective time perception on the injection and post-injection days in the sub–to supra-second temporal range (0.6–2.4 s), but not in the supra-second range (2–8 s).Human and animal studies exploring how acute pain modulates time perception within relatively short temporal ranges have yielded similar results. In humans, lengthened temporal estimation of neutral stimuli has been observed with laser-induced heat pain (0.35–1.37-s stimulus duration) , in a coFew studies have examined the effect of pain on temporal perception over long durations, perhaps because the perceptual process is more complex than for short durations. Subjects must focus their attention on to-be-timed stimuli for a long time, necessitating greater involvement of cognitive abilities such as selective and sustained attention and working memory . In addiPLs to intermediate-duration tones were greater in the formalin group than in the saline group, with no significant difference in the PSE. These results indicate that formalin-induced pain had a weaker effect in this range relative to the 0.6–2.4-s range. In addition, pain is known to automatically attract attention  and did not affect the temporal bisection task on the injection day. Therefore, these less significant changes in saline group may be due to training effects or random fluctuations. Subcutaneous formalin injection into the rat hind paw is a classic method for the creation of a model of acute inflammatory pain. Low-dose  formalin induces phase-I and phase-II nociceptive behaviours that do not last until the following day . HoweverFormalin-induced acute pain did not affect temporal sensitivity in the sub–supra-second or supra-second range in this study. Previous research has revealed a negative relationship between the level of chronic stress and temporal sensitivity  in healthy human subjects . Acute pOur results show that formalin-induced pain affected time perception in the sub- to supra-second range on days after formalin injection, but we conducted observations on only three such days. Thus, the duration of effect persistence remains unclear. Previous studies have shown that animals can learn to be unresponsive to intermediate-duration after repeated performance of the temporal bisection task . To avoiThis study explored the optimal time window for pain to affect time perception, and partially confirmed the possible role of pain emotion in it. These results can provide evidence for some theoretical models of time perception. Striatal beat frequency (SBF) model emphasizes the critical role of cortical activities in temporal information processing . As a muThis study showed that the effect of acute pain on time perception depends on the duration of the tone to be estimated. Acute pain leads to more significant overestimation of shorter-duration than of longer-duration. Pain-induced high arousal and distraction may jointly affect time perception, and this effect may differ between the short-term and long-term ranges. In addition, the emotional component of pain may play a role independent of the sensory component. In future research, the effects of the emotional processing of pain on temporal perception should be explored further.10.7717/peerj.11002/supp-1Data S1Click here for additional data file.10.7717/peerj.11002/supp-2Supplemental Information 1Click here for additional data file.10.7717/peerj.11002/supp-3Figure S1There is no significant difference between formalin group and saline group in each session.Click here for additional data file.10.7717/peerj.11002/supp-4Figure S2p < 0.05, **p < 0.01 vs. saline group.Formalin treatment prolonged response latency to 4-s trials and 5.04-s trials in the injection test (B). *Click here for additional data file.10.7717/peerj.11002/supp-5Figure S3There is no significant difference between formalin group and saline in each session.Click here for additional data file.10.7717/peerj.11002/supp-6Figure S4p < 0.05 vs. saline group.Formalin treatment prolonged the response latency to 4-s trials in post-injection day3 (C). *Click here for additional data file.